title: Expression of a new chimeric protein with a highly repeated sequence in tobacco cells.
abstract: In wheat, the high-molecular weight (HMW) glutenin subunits are known to contribute to gluten viscoelasticity, and show some similarities to elastomeric animal proteins as elastin. When combining the sequence of a glutenin with that of elastin is a way to create new chimeric functional proteins, which could be expressed in plants. The sequence of a glutenin subunit was modified by the insertion of several hydrophobic and elastic motifs derived from elastin (elastin-like peptide, ELP) into the hydrophilic repetitive domain of the glutenin subunit to create a triblock protein, the objective being to improve the mechanical (elastomeric) properties of this wheat storage protein. In this study, we investigated an expression model system to analyze the expression and trafficking of the wild-type HMW glutenin subunit (GS(W)) and an HMW glutenin subunit mutated by the insertion of elastin motifs (GS(M)-ELP). For this purpose, a series of constructs was made to express wild-type subunits and subunits mutated by insertion of elastin motifs in fusion with green fluorescent protein (GFP) in tobacco BY-2 cells. Our results showed for the first time the expression of HMW glutenin fused with GFP in tobacco protoplasts. We also expressed and localized the chimeric protein composed of plant glutenin and animal elastin-like peptides (ELP) in BY-2 protoplasts, and demonstrated its presence in protein body-like structures in the endoplasmic reticulum. This work, therefore, provides a basis for heterologous production of the glutenin-ELP triblock protein to characterize its mechanical properties.
authors.affiliation: Institut National de la Recherche Agronomique, UR1268, Biopolymères Interactions Assemblages, Nantes, France.
authors.forename: Amélie
authors.initials: A
authors.lastname: Saumonneau
authors.forename: Karine
authors.initials: K
authors.lastname: Rottier
authors.forename: Udo
authors.initials: U
authors.lastname: Conrad
authors.forename: Yves
authors.initials: Y
authors.lastname: Popineau
authors.forename: Jacques
authors.initials: J
authors.lastname: Guéguen
authors.forename: Mathilde
authors.initials: M
authors.lastname: Francin-Allami
mesh_terms: D000595:Amino Acid Sequence; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004549:Elastin / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D005983:Glutens / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011523:Protoplasts / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*; D014170:Transformation, Genetic
chemical_list: D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D005983:Glutens; D004549:Elastin; C037302:glutenin